Mechanism of DNA interstrand crosslink repair in vivo

体内DNA链间交联修复机制

• 批准号: 8958561
• 负责人: Justin Courcelle
• 金额: $ 44.55万
• 依托单位: PORTLAND STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-15 至 2019-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8958561
• 关键词: Animal Model; Biochemical Pathway; Biological Assay; Cells; Cellular Assay; Chemicals; Complex; Couples; Coupling; DNA; DNA Damage; DNA Interstrand Crosslinking; DNA Repair; Data; Disease; Escherichia coli; Event; Excision; Genes; Genetic Recombination; Genetic Transcription; Genome; Homologous Gene; Hyperplasia; Lead; Lesion; Malignant Neoplasms; Mammalian Cell; Modeling; Monitor; Mutate; Nature; Nucleotide Excision Repair; Pathway interactions; Process; Proteins; Repair Complex; Replication-Associated Process; Role; Surgical incisions; Work; cancer therapy; crosslink; cytotoxic; effective therapy; endonuclease; global genomic repair; improved; in vivo; mutant; novel therapeutic intervention; nuclease; public health relevance; reconstitution; repaired

 DESCRIPTION (provided by applicant): DNA interstrand crosslinks are a particularly lethal form of DNA damage that represent an absolute block to replication and transcription. Chemicals forming crosslinks have proven to be highly toxic when found in nature, uniquely potent as chemotherapeutics in specific cancers, and effective treatments for a range of diseases states involving hyperplastic or displastic conditions. Although several genes have been isolated that, when mutated, render cells hypersensitive to crosslinks, many aspects of how these complex lesions are repaired and processed in cells remain unknown. Much of what we know about interstrand crosslink repair has come from eukaryotic studies in extracts and suggest that both replication-dependent and replication-independent mechanisms exist. Both mechanisms are proposed to involve multiple repair pathways, coupling components of nucleotide excision repair with recombination, translesion synthesis, as well as other alternative nuclease complexes. However, after the initial incision event, all these models remain highly speculative, and are hampered by the challenges of reconstituting this multi-step, multi-pathway repair process as well as by the complexity and lack of cellular assays available in mammalian cells. Here, we propose to directly identify the cellular pathways and structural intermediates that arise during the repair of interstrand crosslinks in vivo using the model organism of E.coli, where the processes of replication and repair are highly conserved. In E. coli, we have established unique cellular assays to monitor the replication fork processing and global repair for these lesions. In addition, in preliminary data, we show that we have identified an alternative endonuclease, similar to mammalian cells, that couples with the nucleotide excision repair complex and is important for crosslink repair. These assays will allow us to directly and definitively identify the repair and progressive intermediates that arise during crosslink repair i vivo We describe three aims that will be accomplished. 1) We will identify the genes involved in repairing DNA interstrand crosslinks in E. coli and determine whether they operate in a replication-dependent or replication-independent (global genomic) repair pathway in vivo. 2) We will identify the cellular intermediates and biochemical pathway associated with the replication-independent repair of DNA crosslinks in vivo. 3) We will identify the cellular intermediates and biochemical pathway associated with the replication-dependent repair of DNA crosslinks in vivo. The results of these studies will identify the pathways operating in the repair of this medically relevant lesion in vivo and are likely to suggest novel therapeutic approaches that utilize these lesions in the treatment of cancer and other hyperproliferative diseases.

 DNA交联是一种阳离子和转录我们对实习的交联修复的了解是从提取物中的真核研究，并表明复制和两种机制都是由重组，translesion合成以及其他替代络合物涉及多个修复途径IR涉及多个修复途径。多步骤的多条道路修复过程是通过使用大肠杆菌的模型有机体在体内进行的哺乳动物细胞中的复杂性和缺乏细胞分析。复制叉处理和对TESEON的全球修复。 核酸内切酶，类似于哺乳动物细胞，但是核苷酸切除修复的竞争能力将使我们直接并确定确定在Cring Cring Cring Cross链接Tivo期间出现的维修IDIATES，我们描述在大肠杆菌中的交联，并确定它们在依赖性或复制的（全局基因组）修复途径中的作用。 R Intermedites和与体内DNA交叉链接的复制 - 途径相关的生化途径。疾病。

项目成果

Cho Endonuclease Functions during DNA Interstrand Cross-Link Repair in Escherichia coli.

Cho 核酸内切酶在大肠杆菌 DNA 链间交联修复过程中发挥作用。

• DOI: 10.1128/jb.00509-16
• 发表时间: 2016
• 期刊: Journal of bacteriology
• 影响因子: 3.2
• 作者: Perera,AnthonigeVidya;Mendenhall,JamesBrian;Courcelle,CharmainTan;Courcelle,Justin
• 通讯作者: Courcelle,Justin

Replication Rapidly Recovers and Continues in the Presence of Hydroxyurea in Escherichia coli.

在大肠杆菌中存在羟基脲的情况下，复制迅速恢复并继续。

• DOI: 10.1128/jb.00713-17
• 发表时间: 2018
• 期刊: Journal of bacteriology
• 影响因子: 3.2
• 作者: Nazaretyan,SamvelA;Savic,Neda;Sadek,Michael;Hackert,BrandyJ;Courcelle,Justin;Courcelle,CharmainT
• 通讯作者: Courcelle,CharmainT

RecBCD, SbcCD and ExoI process a substrate created by convergent replisomes to complete DNA replication

• DOI: 10.1111/mmi.14242
• 发表时间: 2019-06-01
• 期刊: MOLECULAR MICROBIOLOGY
• 影响因子: 3.6
• 作者: Hamilton, Nicklas A.;Wendel, Brian M.;Courcelle, Justin
• 通讯作者: Courcelle, Justin